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3M Molecular Detection Assay Salmonella Collaborative Study
July 2012
OMA-2012-July-XXX
3.13
Validation Study Confirmation
1
3.13.1
Using the primary enrichment (step 3.1 above), follow steps starting at 4.2
2
below (secondary enrichments) to confirm each test portion, regardless of
3
presumptive result.
4
3.13.2
Transfer typical colonies to TSI/LIA slants. Incubate 35
±
1
°
C for 24
±
5
2h.
6
3.13.3
Confirm a minimum of one typical colony per test portion with
7
biochemical/serological procedures prescribed by the USDA/FSIS
8
method. Either the API20E (Official Method 978.24) or the VITEK GN
9
(Official Method 2011.17) will be used as an alternative to the
10
conventional biochemical tests. The somatic (O) and flagellar (H) tests
11
will also be performed.
12
13
4.0
USDA/FSIS MLG
14
4.1
To each 25g test portion, add 225 mL buffered peptone water. Stomach each
15
sample for approximately two minutes. Incubate at 35
±
1
°
C for 22
±
2h.
16
4.2
Transfer 0.5
±
0.05 mL enriched sample into 10 mL tetrathionate (TT-Hanja)
17
broth and 0.1
±
0.02 mL into 10 mL modified Rappaport-Vassiliadis (mRV)
18
broth.
Pay special attention to the preparation of RV and TT broths as
19
USDA and FDA use different formulations.
Incubate at 42
±
0.5
°
C for 22-24
20
h.
21
4.3
Streak both secondary enrichments onto double modified lysine iron agar
22
(DMLIA) or xylose lysine Tergitol (XLT4) agar and brilliant green sulfa (BGS)
23
agar. Use one loopful of inoculum for each plate. Incubate 35
±
2
°
C for 18-24h.
24
Select colonies. Re-incubate all plates for an additional 18-24 h. Reexamine
25
initially negative plates and pick colonies as above.
26
4.4
Transfer typical colonies to TSI/LIA slants. Incubate 35
±
1
°
C for 24
±
2h.
27
4.5
Confirm a minimum of one typical colony per sample with
28
biochemical/serological procedures prescribed by the USDA/FSIS method. Either
29
the API20E (Official Method 978.24) or the VITEK GN (Official Method
30
2011.17) will be used as an alternative to the conventional biochemical tests. The
31
somatic (O) and flagellar (H) tests will also be performed.
32
33
Final Results
34
35
All results are to be recorded on the data sheets provided. For each test portion, record the
36
results reported by the 3M Molecular Detection Assay
Salmonella
method along with the
37
confirmation results for both the 3M Molecular Detection Assay
Salmonella
and reference
38
methods. Immediately upon completion of each food type, the data sheet should be faxed or
39
emailed to TBD, co-Study Director.
40
DRAFT DOCUMENT
16